We identify and topographically localize inflammatory, degenerative, and malignant cells, as well as their products, in patient specimens and animal tissues. We analyze these cells and their products mainly by routine pathology, immunohistochemistry, and molecular pathology. The application of cutting-edge technology, such as microdissection combined with molecular techniques (PCR, RT-PCR, genotyping, etc,) allows us to provide more accurate pathological diagnosis (assessment) and pathogenesis of the disease. These technologies will guide us in selecting the most targeted treatment for patients. We also learn about the mechanisms of different ocular diseases from various animal models. Using animal models, we can also assess the efficacy of different therapeutic agents for various ocular diseases. In FY2013, we accomplished the following in our research: 1. Molecular Pathology of Age-Related Macular Degeneration (AMD): AMD is the leading cause of irreversible severe central vision loss among the elderly in the world. It is projected that AMD will afflict 3 million Americans over the age of 50 by the year 2020. The pathology of AMD is characterized by the accumulation of soft drusen, RPE and photoreceptor degeneration, geographic atrophy, and/or exudation with choroidal neovascularization in the macula. While several risk factors, including age, race, smoking, and diet have been linked to AMD, the etiology and pathogenesis of the disease remain largely unclear. Treatment options for neovascular AMD include intravitreal injections of anti-vascular endothelial growth factor (VEGF); however, treatment options for geographic atrophy AMD are extremely limited. Current knowledge demonstrates that AMD development is strongly influenced by genetic factors. We and other investigators have reported significant associations between AMD and single nucleotide polymorphisms in CFH, CX3CR1, and ARMS2/HtrA1 over the past years. Growing evidence also suggests that inflammatory and immunological elements (e.g., macrophages and microglia), apoptosis, cholesterol trafficking, angiogenesis (e.g., VEGF expression), and oxidative stress (e.g., the role of mitochondria) play an important role in AMD development. In FY2013, we reviewed and highlighted pathogenic changes that destabilize ocular homeostasis and promote AMD development. With normal aging, photoreceptors are steadily lost, Bruchs membrane thickens, the choroid thins, and hard drusen may form in the periphery. In AMD, many of these changes are exacerbated in addition to the development of disease-specific factors such as soft macular drusen. Para-inflammation, which can be thought of as an intermediate between basal and robust levels of inflammation, develops within the retina in an attempt to maintain ocular homeostasis. Para-inflammation is reflected by increased expression of the anti-inflammatory cytokine IL-10 coupled with shifts in macrophage plasticity from the pro-inflammatory M1 to the anti-inflammatory M2 polarization. In AMD, imbalances in the M1 and M2 populations together with activation of retinal microglia are observed and potentially contribute to tissue degeneration. Nonetheless, the retina persists in a state of chronic inflammation, and increased expression of certain cytokines and inflammasomes is noted. Since not everyone develops AMD, the vital question to ask is how the body establishes a balance between normal age-related changes and the pathological phenotypes in AMD. Aging sets the stage for disease to set in, but the factors that tip the balance in favor of disease should be our targets for therapy. In collaboration with Drs. Lai Wei and Nussenblatt, we identified a significantly decreased level of methylation on the IL17RC promoter in AMD patients. Furthermore, we showed that hypomethylation of the IL17RC promoter in AMD patients led to an elevated expression of its protein and transcript in peripheral blood as well as in the affected macular tissue, suggesting that the DNA methylation pattern and expression of IL17RC may serve as a biomarker for AMD diagnosis and likely plays a role in disease pathogenesis. Using multiple anatomical techniques, we studied the synapses of mice with double deficient Ccl2 and Cx3cr1 on Crb1rd8 background (a murine AMD model) at one month. We demonstrated that cone synapses and opsin, but not rod synapses and rhodopsin, are markedly decreased in focal retinal degenerative lesions in the early phase of photoreceptor degeneration in these mice. We also showed that some rod spheres, but not cone pedicles, form ectopic synapses with postsynaptic processes that express glutamate receptors. These findings could provide new insight to a potential mechanism of vision function loss due to synaptic degeneration before cell death in early AMD. We also found a beneficial effect of the Age-Related Eye Diseases Study 2 (AREDS2) dietary formula containing n-3 polyunsaturated fatty acids, lutein, and zeaxanthin on the retinal AMD-like lesions of these mice. The AREDS2 diet significantly lowered several inflammatory molecules, such as iNOS, COX2, TNF-&#945;, and IL-1&#946;, as well as VEGF in sera and/or retina. Although the findings in a homogeneous mouse population may not necessarily translate to findings in humans, they could still provide useful information for our understanding AMD pathogenesis and investigating novel therapeutic targets. 2. Ocular Lymphoma: We summarized the current concepts in diagnosing and managing of primary vitreoretinal lymphoma (PVRL). PVRL, previously called primary intraocular lymphoma (PIOL), is a rare and fatal ocular malignancy. The diagnosis of PVRL is often challenging as it often masquerades as chronic uveitis. PVRL requires invasive procedures for tissue diagnosis. Cytology/pathology, molecular pathology, immunohistochemistry, flow cytometry, and cytokine analysis (IL-10) are often needed. Current treatment of PVRL involves aggressive chemotherapy and radiation therapy. A professional team of medical experts in ophthalmology, oncology (neurooncology), and pathology is essential for optimizing patient management. We published our findings on the association between IL-10-1082 SNP and PVRL / PCNSL. More PVRL patients expressed one copy of the IL-10-1082 SNP genotype compared to controls. The IL-10-1082 A allele is a risk factor for higher IL-10 levels in the vitreous of patients with PVRLs and PCNSLs. Higher IL-10 levels have been correlated with more aggressive disease in both PVRLs and PCNSLs, making this finding an important and potentially clinically significant observation. We reported another case of PVRL and testicular lymphoma and showed that lymphoma in an immune-privileged site such as the testis may recur in the eye, warranting careful surveillance. We also reported that anti-LFA-1 antibodies enhance metastasis of ocular lymphoma to the brain and contralateral eye in a murine model of intraocular lymphoma. The findings suggest that confinement of lymphoma cells to the eye depends on active immune surveillance using a population of effector cells expressing the cell surface integrin LFA-1. 3. New Pathology and Pathogenesis of Ocular Diseases: In collaboration with Dr. Singh of Cleveland Clinic, we presented ultrastructure and immunoreactivity of the eyes of a patient with melanoma and paraneoplastic vitelliform retinopathy. We provided convincing histological evidence of melanoma-associated autoantibodies acting directly against transient receptor potential M1 channels (TRPM1) that target the ON bipolar cell structure in the inner nuclear and outer plexiform layers in paraneoplastic vitelliform retinopathy. In collaboration with Dr. Catherine Morgans of Oregon Health and Sciences University and Dr. Nida Sen of NEI, we reported the first case of melanoma-associate autoimmune retinopathy.